Sci. Aging Knowl. Environ., 26 February 2003
Bridge Inside Troubled Cells
Stress unites protective proteins by forming atomic ties
Key Words: redox regulation Hsp70 Hsp27 monomer transcription
Smoke across the street from a firehouse can rouse the rescue squad without the ring of an emergency phone call. Similarly, heat and oxidative stress directly motivate a protective protein, rather than acting through intermediate steps to save the cell, according to new work. The results illuminate the molecular details of how environmental insults incite cellular defense systems.
Abating stress is one secret to long life. From yeast to humans, cells employ a protein called heat shock factor-1 (hsf-1) to engage defense systems that guard against life-threatening assaults. High temperatures and oxidative stress power up hsf-1 molecules by prompting them to band together in groups of three. These trimers bind to particular DNA sequences and turn on nearby genes whose protein products help clean up the cellular mess. Researchers have long wondered how stressors provoke mammalian hsf-1 to unite.
To address the issue, Ahn and Thiele incubated hsf-1 in a test tube at high temperature or in the presence of hydrogen peroxide, a chemical that inflicts oxidative damage. After either treatment, the individual proteins coalesced into threesomes, revealing for the first time that the stresses act directly on hsf-1 and not through cellular go-betweens. Then, the researchers sought the chemical changes that make the proteins stick together. Disulfide bonds--chemical bridges between sulfur atoms--help many proteins maintain their shape; oxidizing conditions--such as those imposed by hydrogen peroxide--favor the formation of such bridges. Because hsf-1 carries the sulfur-containing amino acid cysteine at several locations in its protein sequence, the investigators wondered whether cysteine couplings stabilize hsf-1 trimers. To test the idea, they used heat or oxidative stress to generate hsf-1 triplets in a test tube. Then, they added a chemical that severs cysteine connections, and the protein clusters fell apart. Removing the agent and again stressing the protein caused hsf-1 to regroup. The results suggest that the formation of hsf-1 trimers depends on intact disulfide bonds.
To verify the importance of the cysteine spans, the researchers created mutant hsf-1 molecules in which they replaced a cysteine with a different amino acid incapable of forming bridges. When subjected to heat or oxidative stress, the mutants could not associate into trimers. In addition, the altered proteins could not perform hsf-1's normal tasks in live cells, such as entering the nucleus and turning on appropriate genes when stressed. Taken together, the results suggest that heat and oxidative stress regulate hsf-1 in the cell by spurring cysteines to connect.
The finding is "very significant," says molecular biologist Ivor Benjamin of the University of Texas Southwestern Medical Center in Dallas. Researchers who study the heat shock response have been grappling with how it is regulated, and this study sheds new light on that issue, he says--knowledge that might help researchers keep stress from burning up years.
February 26, 2003
Science of Aging Knowledge Environment. ISSN 1539-6150